Publications

Mechanical changes in tumors have long been linked to increased malignancy and therapy resistance and attributed to mechanical changes in the tumor extracellular matrix (ECM). However, to the best of our knowledge, there have been no mechanical studies on decellularized tumors. Here, we studied the biochemical and mechanical progression of the tumor ECM in two models of lung metastases: lung carcinoma (CAR) and melanoma (MEL). We decellularized the metastatic lung sections, measured the micromechanics of the tumor ECM, and stained the sections for ECM proteins, proliferation, and cell death markers. The same methodology was applied to MEL mice treated with the clinically approved anti-fibrotic drug nintedanib. When compared to healthy ECM (~0.40 kPa), CAR and MEL lung macrometastases produced a highly dense and stiff ECM (1.79 ± 1.32 kPa, CAR and 6.39 ± 3.37 kPa, MEL). Fibronectin was overexpressed from the early stages (~118%) to developed macrometastases (~260%) in both models. Surprisingly, nintedanib caused a 4-fold increase in ECM-occupied tumor area (5.1 ± 1.6% to 18.6 ± 8.9%) and a 2-fold in-crease in ECM stiffness (6.39 ± 3.37 kPa to 12.35 ± 5.74 kPa). This increase in stiffness strongly correlated with an increase in necrosis, which reveals a potential link between tumor hypoxia and ECM deposition and stiffness. Our findings highlight fibronectin and tumor ECM mechanics as attractive targets in cancer therapy and support the need to identify new anti-fibrotic drugs to abrogate aberrant ECM mechanics in metastases.

JTD Keywords: atomic force microscopy, basement membrane, breast-cancer, decellularization, expression, extracellular matrix, extracellular-matrix, fibronectin, intermittent hypoxia, lung carcinoma, lung metastases, melanoma, metastatic niche formation, micromechanical properties, nintedanib, signature, stiffness, tumor-growth, Colorectal-cancer progression, Lung metastases, Stiffness

Objective: Intermittent hypoxia (IH)—a hallmark of obstructive sleep apnea (OSA)—enhances lung cancer progression in mice via altered host immune responses that are also age and sex-dependent. However, the interactions of menopause with IH on tumor malignant properties remain unexplored. Here, we aimed to investigate lung cancer outcomes in the context of ovariectomy (OVX)-induced menopause in a murine model of OSA. Methods: Thirty-four female mice (C57BL/6, 12-week-old) were subjected to bilateral OVX or to Sham intervention. Six months after surgery, mice were pre-exposed to either IH or room air (RA) for 2 weeks. Then, 105 lung carcinoma (LLC1) cells were injected subcutaneously in the left flank, with IH or RA exposures continued for 4 weeks. Tumor weight, tumor invasion, and spontaneous lung metastases were assessed. Tumor-associated macrophages (TAMs) were isolated and subjected to flow cytometry polarity evaluation along with assessment of TAMs modulation of LLC1 proliferation in vitro. To determine the effect of IH and OVX on each experimental variable, a two-way analysis of variance was performed. Results: IH and OVX promoted a similar increase in tumor growth (2-fold; P = 0.05 and 1.74-fold; P < 0.05, respectively), and OVX-IH further increased it. Regarding lung metastasis, the concurrence of OVX in mice exposed to IH enhanced the number of metastases (23.7 ± 8.0) in comparison to those without OVX (7.9 ± 2.8; P < 0.05). The pro-tumoral phenotype of TAMS, assessed as M2/M1 ratio, was increased in OVX (0.06 ± 0.01; P < 0.01) and IH (0.06 ± 0.01; P < 0.01) compared with sham/RA conditions (0.14 ± 0.03). The co-culture of TAMS with naive LLC1 cells enhanced their proliferation only under IH. Conclusion: In female mice, both the IH that is characteristically present in OSA and OVX as a menopause model emerge as independent contributors that promote lung cancer aggressiveness and seemingly operate through alterations in the host immune response.

JTD Keywords: Animal models, Cancer progression, Intermittent hypoxia, Menopause, Obstructive sleep apnea, Ovariectomy